To Heat Treat or Not to Heat Treat Calf Serum, that is the Question?

Ok, so the Bard is probably spinning in his grave at our blatant plagiarism of the script from the “Scottish Play” (it is bad luck to use its real name, apparently).  However, it seems an apt way to introduce the question of whether or not you should heat treat foetal bovine serum (FBS) before using it to propagate insect cells.  Of course, Spodoptera or Trichoplusia spp. cells are mostly grown in serum-free media so the problem doesn’t arise.  For S. frugiperda (Sf21) cells or T. ni TN368 lines these are usually grown in serum-containing medium (normally based on TC100).  Although their usage is far less than cells grown in serum-free media, they remain an important tool for certain applications, such as plaque assay-based titration of recombinant virus in Sf21 cells.  FBS is also often added to recombinant virus stocks previously propagated in cells grown in serum-free medium to aid their preservation.

Historically, calf sera have been heat treated (56°C for 30min) prior to use with media for propagating mammalian cell lines.  The rationale for this is shrouded in uncertainty.  Traditionally, the reason for heat treatment was to inactivate residual complement in the serum, which might affect cell growth.  However, levels of complement in FBS have been shown to be very low (Triglia and Linscott [1980] Journal of Mol. Immunol., vol. 17, pp 741-748, 1980).  Furthermore, complement might not be expected to affect insect cells.  Despite this, on occasion we have found that non-heat inactivated FBS used at 10% (v/v) with TC100 can have an adverse effect on cell growth.  This problem came to the fore just before Christmas when we tested a new batch of FBS from a commercial supplier.  Tests with and without FBS revealed that Sf21 cells performed best when grown in TC100 with 10% heat inactivated FBS.  Non heat-treated serum did support cell growth, but it was definitely sub-optimal.

The above observation serves to emphasize that new batches of FBS should be tested for compatibility with insect cells.  Serum samples should be heat-treated or left in their original condition and then used to maintain insect cells for at least two passages.  If similar growth rates are observed with each FBS sample then there is no need to heat-treat the serum.  Depending on your budget, you can then purchase an amount of FBS sufficient to keep you supplied for at least a year to avoid repeating this test too frequently.